Topic 1a Flashcards
Cell
- carries hereditary information that defines the species
- capable of self replication
Discovery of Cells
- Z & H Janssen
first microscope - Robert Hooke
coined term “cell” after describing chambers in cork - Anton van Leenwenhoek
made his own refined microscopes (300x); first to observe living cells (animacules) in pond water and tooth scrapings
Birth of Modern Cell Biology
- Schleden and Schwann introduced first 2 rules of the Cell Theory
- all life is made from 1+ cells
- cell is basic unit of structure for all organisms - Virchow added the final rules
- all cells arise from pre-existing cells
Schleden and Schwann
Introduced first 2 rules of the Cell Theory
- all life is made from 1+ cells
- cell is basic unit of structure for all organisms
Virchow
Modified Schleden and Shwann’s Cell Theory
- all cells arise from pre-existing cells
Craig Venter
participated in human genome project
created first artificially created cell
Development of Cell Biology
- Cytology: emerged with Van Leewenhoek’s improvements upon the microscope.
- Biochemistry: facilitated by advancements in bioogical tecniques
- Genetics
Micrometer (um)
One-millionth of a meter, 1/25000 of an inch.
- unit of choice for describing cells and organelles
Nanometer
Unit of choice for molecules and subcellular structures; 1/billionth of a meter.
Microtome
Instrument developed for the rapid and efficient preparation of thin tissue slices.
Limit of Resolution & Resolving Power
LR: A quality refering to how far apart adjacent objects must be to appear as seperate entities.
RP: A microscope quality refering to the ability to see fine details of structure
- the smaller the LR, the greater (&better!) the RP
- using a light microscope under violet light, the theoretical LR is around 200nm
Brightfield Microscopy
White light is passed directly through a stained or unstained specimen while the field (background) is illuminated.
-limitation: specimen must be unalive and specifcally prepared to highlight transparent features
Types of Microscopy
- Phase contrast: enhances contrast in unstained cells by amplifying variations in refractive index within specimen; especially useful for examining living, unpigmented cells.
- Differential Interference Contrast: also uses optical modifications to exaggerate differences in refractive index
- Fluorescence: Shows locations of specific molecules within cell. Fluorescent substances absorb ultraviolet radiation and emit visible
light. The fluorescing molecules may occur naturally but more often are made by tagging the molecules of interest with fluorescent dyes or antibodies. - Confocal: Uses lasers and special optics to focus illuminating beam on a single plane within the specimen. Only those regions within a narrow depth of focus are imaged. Regions above and below the selected plane of view appear black rather than blurry.
- Brightfield: passes light directly through specimen; image has little contrast unless specimen is stained
Phase Contrast Microscopy
Enhances contrast in unstained cells by amplifying variations in refractive index within specimen; especially useful for examining living, unpigmented cells
- breaks waves which are in phase as a sample of light hits a specimen, then brings them back together at the eyepiece
Fluorescence Microscopy
Shows locations of specific molecules within cell. Fluorescent substances absorb ultraviolet radiation (short wavelength) and emit visible light (long wavelength)
- utilizes DICHROMAIC MIRROR; reflects short wavelength and transmits longer wavelengths
- fluorescing molecules may occur naturally but more often are made by tagging the molecules of interest with fluorescent dyes or antibodies.
- ideal wavelength for filter must be inbetween absorbance and reflective cutoffs (ie. abs. at 470, ref. at 550; dichoric mirror ideal at 470)
- resolution is not increased, you can just see more because it is illuminated
Differential Interference Contrast/ Nomarsky Microscopy
uses optical modifications to exaggerate differences in refractive index; good for living cells
Confocal Microscopy and Z-stacks
Specimen image is scanned on multiple focal plates and images are stacked upon eachother (z-stacks)to create a 3D image
Antibody
Protien molecule produced by the immune system which binds to one target molecule/antigen
- allows microscopers to visualize and identify soecific molecules within cells
Angstrom (Å)
0.1 nm, size of a hydrogen atom
Sizes of Cells & their Components
Typical eukaryotic cell: 10-100um
Typical prokaryotic cell: 1-10um
Light microscope limit of res: 0.2nm
Electron microscope limit of res: 0.2 nm/ 2 Å
Resolution Equation
resolution = 0.61 λ /NA
λ = wavelength (nm)
NA = numerical aperture = nsinθ
- the smaller the better!
Importance of Wavelength
We are limited by the range of violet light (390nm), which is too large to hit small objects under a microscope; electromagnetic waves (0.004nm), allows for greater visibility.
Digital Video Microscopy
attatches light-sensitive video camera to a light microscope to collect digital images for storage.
-allows cells to be observed for extended period of time
Immunofluorescence
A fluorescent molecule is attached to an antibody, which in turn recognizes and binds to one specific complementary target molecule/ antigen
- for fluorescence or confocal microscope